Ellur Govindaraj, Govindappa Prem Kumar, Subrahmanian Sandeep, Romero Gerardo Figueroa, Gonzales David A, Margolis David S, Elfar John C
Department of Orthopaedics and Sports Medicine, University of Arizona College of Medicine, Tucson, Arizona.
Penn State Milton S. Hershey Medical Center, Hershey, Pennsylvania.
J Bone Joint Surg Am. 2025 May 7;107(9):936-947. doi: 10.2106/JBJS.24.00311. Epub 2025 Mar 26.
Delayed bone healing is common in orthopaedic clinical care. Agents that alter cell function to enhance healing would change treatment paradigms. 4-aminopyridine (4-AP) is a U.S. Food and Drug Administration (FDA)-approved drug shown to improve walking in patients with chronic neurological disorders. We recently showed 4-AP's positive effects in the setting of nerve, wound, and even combined multi-tissue limb injury. Here, we directly investigated the effects of 4-AP on bone fracture healing, where differentiation of mesenchymal stem cells into osteoblasts is crucial.
All animal experiments conformed to the protocols approved by the Institutional Animal Care and Use Committee at the University of Arizona and Pennsylvania State University. Ten-week-old C57BL/6J male mice (22 to 28 g), following midshaft tibial fracture, were assigned to 4-AP (1.6 mg/kg/day, intraperitoneal [IP]) and saline solution (0.1 mL/mouse/day, IP) treatment groups. Tibiae were harvested on day 21 for micro-computed tomography (CT), 3-point bending tests, and histomorphological analyses. 4-AP's effect on human bone marrow mesenchymal stem cell (hBMSC) and human osteoblast (hOB) cell viability, migration, and proliferation; collagen deposition; matrix mineralization; and bone-forming gene/protein expression analyses was assessed.
4-AP significantly upregulated BMP2 gene and protein expression and gene expression of RUNX2, OSX, BSP, OCN, and OPN in hBMSCs and hOBs. 4-AP significantly enhanced osteoblast migration and proliferation, collagen deposition, and matrix mineralization. Radiographic and micro-CT imaging confirmed 4-AP's benefit versus saline solution treatment in mouse tibial fracture healing (bone mineral density, 687.12 versus 488.29 mg hydroxyapatite/cm 3 [p ≤ 0.0021]; bone volume/tissue volume, 0.87 versus 0.72 [p ≤ 0.05]; trabecular number, 7.50 versus 5.78/mm [p ≤ 0.05]; and trabecular thickness, 0.08 versus 0.06 mm [p ≤ 0.05]). Three-point bending tests demonstrated 4-AP's improvement of tibial fracture biomechanical properties versus saline solution (stiffness, 27.93 versus 14.30 N/mm; p ≤ 0.05). 4-AP also increased endogenous BMP2 expression and matrix components in healing callus.
4-AP increased the healing rate, biomechanical properties, and endogenous BMP2 expression of tibiae following fracture.
Prognostic Level III . See Instructions for Authors for a complete description of levels of evidence.
在骨科临床治疗中，骨愈合延迟很常见。能够改变细胞功能以促进愈合的药物将改变治疗模式。4-氨基吡啶（4-AP）是一种经美国食品药品监督管理局（FDA）批准的药物，已证明可改善慢性神经疾病患者的行走能力。我们最近发现4-AP在神经、伤口甚至多组织肢体联合损伤的情况下具有积极作用。在此，我们直接研究了4-AP对骨折愈合的影响，其中间充质干细胞向成骨细胞的分化至关重要。
所有动物实验均符合亚利桑那大学和宾夕法尼亚州立大学机构动物护理与使用委员会批准的方案。10周龄的C57BL/6J雄性小鼠（22至28克），在胫骨中段骨折后，被分配到4-AP（1.6毫克/千克/天，腹腔注射[IP]）和生理盐水（0.1毫升/小鼠/天，腹腔注射）治疗组。在第21天采集胫骨进行微型计算机断层扫描（CT）、三点弯曲试验和组织形态学分析。评估了4-AP对人骨髓间充质干细胞（hBMSC）和人成骨细胞（hOB）的细胞活力、迁移、增殖；胶原蛋白沉积；基质矿化；以及骨形成基因/蛋白表达分析。
4-AP显著上调了hBMSC和hOB中BMP2基因和蛋白表达以及RUNX2、OSX、BSP、OCN和OPN的基因表达。4-AP显著增强了成骨细胞的迁移和增殖、胶原蛋白沉积以及基质矿化。影像学和微型CT成像证实了4-AP在小鼠胫骨骨折愈合方面相对于生理盐水治疗的益处（骨矿物质密度，687.12对488.29毫克羟基磷灰石/立方厘米[p≤0.0021]；骨体积/组织体积，0.87对0.72[p≤0.05]；骨小梁数量，7.50对5.78/毫米[p≤0.05]；以及骨小梁厚度，0.08对0.06毫米[p≤0.05]）。三点弯曲试验表明4-AP相对于生理盐水改善了胫骨骨折的生物力学性能（刚度，27.93对14.30牛/毫米；p≤0.05）。4-AP还增加了愈合骨痂中内源性BMP2的表达和基质成分。
4-AP提高了骨折后胫骨的愈合率、生物力学性能和内源性BMP2表达。
预后水平III。有关证据水平的完整描述，请参阅作者须知。
